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Targeted metabolomics analysis of Campylobacter coli VC167 reveals legionaminic acid derivatives as novel flagellar glycans

From National Research Council Canada

DOIResolve DOI: https://doi.org/10.1074/jbc.M611027200
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Affiliation
  1. National Research Council of Canada. NRC Institute for Biological Sciences
  2. National Research Council of Canada. NRC Institute for Marine Biosciences
FormatText, Article
Subjectbiosynthetic pathways; Campylobacter coli; Campylobacter jejuni; Canada; chemistry; chromatography, liquid; complement; cyclic amp; enzymes; flagellin; gene expression regulation, bacterial; genes, bacterial; genetics; glycans; glycosylation; interaction; ions; magnetic resonance spectroscopy; mass spectrometry; metabolism; models, chemical; models, molecular; molecular sequence data; mutagenesis; mutants; mutation; NMR spectroscopy; nucleotides; pathway; polysaccharides; protein; role; sialic acids; spectrometry, mass, matrix-assisted laser desorption-ionization; spectroscopy; structural analysis; sugars
Abstract
Glycosylation of Campylobacter flagellin is required for the biogenesis of a functional flagella filament. Recently, we used a targeted metabolomics approach using mass spectrometry and NMR to identify changes in the metabolic profile of wild type and mutants in the flagellar glycosylation locus, characterize novel metabolites, and assign function to genes to define the pseudaminic acid biosynthetic pathway in Campylobacter jejuni 81-176 (McNally, D. J., Hui, J. P., Aubry, A. J., Mui, K. K., Guerry, P., Brisson, J. R., Logan, S. M., and Soo, E. C. (2006) J. Biol. Chem. 281, 18489-18498). In this study, we use a similar approach to further define the glycome and metabolomic complement of nucleotide-activated sugars in Campylobacter coli VC167. Herein we demonstrate that, in addition to CMP-pseudaminic acid, C. coli VC167 also produces two structurally distinct nucleotide-activated nonulosonate sugars that were observed as negative ions at m/z 637 and m/z 651 (CMP-315 and CMP-329). Hydrophilic interaction liquid chromatography-mass spectrometry yielded suitable amounts of the pure sugar nucleotides for NMR spectroscopy using a cold probe. Structural analysis in conjunction with molecular modeling identified the sugar moieties as acetamidino and N-methylacetimidoyl derivatives of legionaminic acid (Leg5Am7Ac and Leg5AmNMe7Ac). Targeted metabolomic analyses of isogenic mutants established a role for the ptmA-F genes and defined two new ptm genes in this locus as legionaminic acid biosynthetic enzymes. This is the first report of legionaminic acid in Campylobacter sp. and the first report of legionaminic acid derivatives as modifications on a protein
Publication date
PublisherAmerican Society for Biochemistry and Molecular Biology
In
  • The Journal of Biological Chemistry 282, no. 19: 14463–14475.
LanguageEnglish
Peer reviewedYes
NRC numberMCNALLY2007A
1702
NPARC number9362866
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Record identifierbc8fa29a-3a30-4ae1-94ef-2153a3dd23bb
Record created2009-07-10
Record modified2020-05-10
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